Summary: Since both humoral and cellular and immune responses may be important against AIDS, two HIV-1 multigenic plasmid DNAs were constructed containing the gag and env genes from pNL4-3 DNA under the regulation of the human CMV and murine endogenous AKV promoters. Development of humoral and T cell proliferative immune responses were analyzed by direct DNA injection of rhesus and pig-tailed macaques. Similar levels of T-cell proliferative responses were generated with both DNAs in rhesus macaques whereas the humoral responses in rhesus and pig-tailed macaques correlated directly with the promoter strength of the vaccine DNA. Long-term, boostable antibodies to Gag and Env were generated using less amount and fewer injections of CMV-DNA than with AKV-DNA. Tetanus toxoid injection was used to demonstrate ability of the animals to respond similarly to a known antigen. To evaluate protection against HIV-1, DNA vaccinated pig-tailed and rhesus macaques were boosted with a single injection of oligomeric gp160 (to enhance the env response). Evaluation of the Env antibody responses at 4 weeks post-boost indicated increased antibody titers to gp120 and gp41 in the HIV-1 DNA vaccinated animal as compared with the control animals. No neutralizing antibodies were detected. The rhesus macaques were challenged with SHIV-IIIb. At 4 weeks post-protein boost, the HIV-1 DNA vacine + protein immunization resulted in reduction of plasma viral RNA indicating that the animals were partially protected. Pig-tailed macaques were challenged with HIV-1. Plasma viral load analysis of the HIV-1 DNA vaccinated animals demonstrated absence of detectable virus by bDNA PCR in animals vaccinated with the CMV-HIV DNA. Studies are underway to investigate viral sequences in the PBMCs to determine whether complete protection was achieved in the DNA vaccinated animals.